Mistletoe (Viscum album) has been known from antiquity as a healing plant. The semishrub plant lives as a semiparasite on the branches of woody plants and is particularly widespread in Europe, North Australia, Asia and in tropical and subtropical Africa. At the start of this century, the cyto- and tumour-toxic action of mistletoe extract, which has since then been specifically used for cancer therapy, was recognised. For this, the extract is used both as a single therapeutic agent and also in combination with chemo- or radiation therapy. Mistletoe preparations are particularly often used for example as a prophylactic against relapse after surgical tumour removal.
Systematic studies of the mode of action show that, after injection, aqueous mistletoe extract as well as its cytotoxic action also has an immunomodulatory effect, and apart from this shows generally mood-brightening effects. After injection of mistletoe extract, a significant increase in the cell numbers of certain lymphocyte subpopulations (inter alia T helper lymphocytes, natural killer (NK) cells and macrophages) and phagocytosis activity in granulo- and monocytes, which are directly involved in tumour defense, are observed (Hajto T, Hostanska K, Gabius H-J, (1990), Therapeutikum 4, 135-145; Beuth J, Ko H L, Tunggal L, Gabius H-J, Steuer M, Uhlenbruck G, Pulverer G (1993), Med. Welt 44, 217-220; Beuth J, Ko H L, Tunggal L, Geisel J, Pulverer G (1993), Arzneim.-Forsch/Drug Res. 43 (1), 166-169; Beuth J, Ko H L, Gabius H-J, Burricheter H, Oette Kl, Pulverer G (1992), Clin. Investing, 70, 658-661). Further, a significant increase in defined acute phase proteins in the serum, which is mediated by the cytokines IL-1, IL-6 and TNF-α, can be detected (Hajto T, Hostanska K, Frei K, Rordorf C, Gabius H-J (1990), Cancer Res. 50, 3322-3326; Beuth J, Ko H-L, Gabius H-J, Pulverer G (1991), In Vivo 5, 29-32; Beuth J, Ko H-L, Tunggal L, Jeljaszewicz J, Steuer M K, Pulverer G (1994), In Vivo 8, 989-992; Beuth J, Ko H-L, Tunggal L, Jeljaszewicz J, Steuer M K, Pulverer G (1994), Dtsch. Zschr. Onkol. 26, 1-6; Beuth J, Ko H-L, Tunggal L, Steuer M K, Geisel J, Jeljaszewicz J, Pulverer G (1993), In Vivo 7, 407-410, Kayser K, Gabius S, Gabius H-J, Hagemeyer O (1992) Tumordiag. und Ther. 13, 190-195). As well as the prolongation of the survival time of cancer patients achievable by mistletoe extract treatment, an increase in the patients' quality of life is also observed, which is attributed to the rise in β-endorphins in the blood (Heiny B-M, Beuth J (1994), Anticancer Res. 14, 1339-1342; Heiny B-M, Beuth J (1994), Dtsch. Zschr. Onkol. 26, 103-108). As endogenous opioids, β-endorphins improve the general well-being, in that they for example have a pain-relieving action, and improve the pain index (Falconer J, Chan E C, Madsens G (1988), J. Endocrinol. 118, 5-8).
Analysis of the active substances of mistletoe extract has shown that the immunostimulating effect is attributable to a certain group of glycoproteins, the mistletoe lectins. Hitherto, three mistletoe-specific lectins with different molecular weights and sugar-binding specificities had been identified. The concentration of mistletoe lectin I (ML-I) in the aqueous plant extract is markedly higher than that of mistletoe lectin II (ML-II) and mistletoe lectin III (ML-III). It could be shown that the immunostimulating effect of the mistletoe extract is attributable to the presence of ML-I: if the ML-I lectin is removed from the mistletoe extract, the extract loses its immunostimulating action (Beuth J, Stoffel B, Ko H-L, Jeljaszewicz J, Pulverer G (1995), Arzneim.-Forsch./Drug. Res. 45 (II), 1240-1242). The β-galactoside-specific ML-I lectin consists of two A- and two B-chains (MLA and MLB), each glycosylated, whose molecular weights are about 29 kDa and 34 kDa respectively. The amino acid sequence of MLA contains one potential glycosylation site, while MLB contains three glycosylation sites in the N-terminal region of the amino acid sequence. The two chains are linked together via a disulphide bridge (FIG. A; Ziska P, Franz H, Kindt A (1978), Experientia 34, 123-124). The resulting mistletoe lectin monomers can associate into dimers with the formation of non-covalent bonds.
Studies of the sedimentation behaviour of ML-I during analytical centrifugation show that in vivo ML-I is present in a monomer-dimer equilibrium (Luther P, Theise H. Chatterjee B, Kardruck D, Uhlenbruck G (1980), Int. J. Biochem. 11, 429-435). The MLB-chain is able to bind to galactose-containing structures on the surface of cell membranes (e.g. receptor molecules) and thereby to trigger cytokine release. Through endocytosis, ML-I dimers and monomers get into the cell, where the protein complexes break down into MLA and MLB chains through reduction of the disulphide bridge bonds. The MLA chains are thereupon able to bind to the ribosomal 28 S subunit and to inactivate this.
The study of ML-I monomers using 2-D gel electrophoresis yielded 25 different isoforms, which are attributable to different combinations of various A and B chains and different glycosylation states of the chains (Schink et al., 1992, Naturwissenschaften 79, 80-81). It is suspected that the individual isoforms fulfil specific functions and each of these isoforms contributes to the anti-tumorigenic effect of the mistletoe extract.
By now, a nucleic acid sequence and the amino acid sequence derived therefrom of one ML-I lectin is already known from European Patent Application EP 0 751 211 A1. However this one polypeptide is not capable of satisfactorily emulating the action of the many ML-I isoenzymes contained in natural mistletoe extract as regards the anti-tumorigenic and mood-brightening effect.